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United States Patent |
6,166,230
|
Bijl
,   et al.
|
December 26, 2000
|
Sterol extraction with polar solvent to give low sterol, high
triglyceride, microbial oil
Abstract
The present invention relates to a process of treating an oil, the process
comprising contacting the oil with a polar solvent to extract at least one
compound that is soluble in the solvent, and then separating the solvent
containing the compound from the so treated oil. The oil is microbially
derived, and extracted either from a fermentation broth or a filtrate
thereof using hexane. The compound to be extracted is usually a sterol or
a diglyceride. The solvent is ethanol having up to 5% water. The oil can
contain a polyunsaturated fatty acid such as C18, C20 or C22 .omega.-3 or
.omega.-6 fatty acid, such as arachidonic acid.
Inventors:
|
Bijl; Hendrik Louis (Vlaardingen, NL);
Wolf; Johannes Hendrik (Delft, NL);
Schaap; Albert (Barendrecht, NL)
|
Assignee:
|
Gist-brocades B.V. (Ma Delft, NL)
|
Appl. No.:
|
180780 |
Filed:
|
February 8, 1999 |
PCT Filed:
|
May 15, 1997
|
PCT NO:
|
PCT/EP97/02510
|
371 Date:
|
February 8, 1999
|
102(e) Date:
|
February 8, 1999
|
PCT PUB.NO.:
|
WO97/43362 |
PCT PUB. Date:
|
November 20, 1997 |
Foreign Application Priority Data
Current U.S. Class: |
554/1 |
Intern'l Class: |
C07C 053/00 |
Field of Search: |
554/8,20,206
435/134
|
References Cited
Foreign Patent Documents |
86/04354 | Jul., 1986 | WO | .
|
Other References
Japan Abstr. of JP62/65689, 1987.
Derwent Abstr. of JP62/798598, 1987.
Yamada, "Production of Dihomo-.gamma.-linolenic Acid, Arachidonic Acid and
Eicosapentaenoic Acid by Filamentous Fungi", Industrial Applications of
Single Cell Oils, Eds. Kyle and Rathledge, (1992) 118-138.
|
Primary Examiner: Carr; Deborah D.
Attorney, Agent or Firm: Morrison & Foerster, LLP
Parent Case Text
This application is a 371 of PCT/EP97/02510 filed May 5, 1997.
Claims
What is claimed is:
1. A process of treating an oil derived from a microorganism, the process
comprising:
(a) contacting the oil with a polar solvent to extract at least one sterol
that is soluble in the solvent; and
(b) separating at least some of the solvent containing the sterol from the
oil, wherein the oil has a sterol content of less than 1.5%.
2. A process according to claim 1 when the oil is obtained or extracted
from a composition resulting from a fermentation.
3. A process according to claim 2 when the composition is a fermentation
broth.
4. A process according to claim 2 wherein the oil is derived, obtained or
extracted from microorganisms present in the composition.
5. A process according to claim 4 wherein the microorganisms are first
removed from the composition.
6. A process according to claim 5 wherein the microorganisms are first
removed by filtering the composition.
7. A process according to claim 5 wherein the microorganisms are dried
before the oil is obtained.
8. A process according to claim 2 wherein the oil has been extracted using
a solvent for triglycerides.
9. A process according to claim 8 when the solvent is hexane, supercritical
carbon dioxide or isopropanol.
10. A process according to claim 1 wherein the oil is produced by, or the
microorganisms is, a bacterium, fungus, yeast or alga.
11. A process according to claim 10 when the microorganism is of the genus
Crypthecodinium, Mucorales, Thraustochytrium, Mortierella, Pythium,
Entomophthora, Porphyridium or Nitzschia.
12. A process according to claim 1 wherein the oil is derived from
Mortierella alpina.
13. A process according to claim 1 wherein the sterol is produced by, or is
present intracellularly inside, the microorganism.
14. A process according to claim 1 wherein the sterol is desmosterol.
15. A process according to claim 1 when the oil comprises at least one
polyunsaturated fatty acid (PUFA).
16. A process according to claim 15 herein the PUFA is a C18, C20 or C22
.omega.-3 or .omega.-6 polyunsaturated fatty acid.
17. A process according to claim 16 wherein the PUFA is GLA, DLA, ARA, EPA
or DHA.
18. A process for preparing an oil comprising at least one polyunsaturated
fatty acid (PUFA), the process comprising:
(a) treating an oil derived from a microorganism and comprising at least
one PUFA and at least one sterol with a polar solvent to extract at least
some of the PUFA and at least some of the sterol, both the PUFA and sterol
being at least partially soluble in the solvent; and
(b) separating the solvent from the oil, and evaporating or otherwise
removing some of the solvent,
to give a (residual) oil having a sterol content of at least 10%.
19. A process according to claim 18 when the sterol is desmosterol.
20. A process according to claim 18 wherein the PUFA is a C18, C20 or C22
.omega.-3 or .omega.-6 polyunsaturated fatty acid.
21. A process according to claim 20 when the PUFA is GLA, DLA, ARA, EPA or
DHA.
22. A process according to claim 1 wherein the solvent comprises a
C.sub.1-6 alkanol or acetone.
23. A process according to claim 22 wherein the solvent is ethanol or
isopropanol.
24. A process according to claim 1 wherein the solvent comprises ethanol
and from 1% to 5% water.
25. A process according to claim 1 wherein the amount of solvent used in
the extraction is from 1 to 9 times the volume of the oil to be treated.
26. A process according to claim 18 wherein the solvent comprises a
C.sub.1-6 alkanol or acetone.
27. A process according to claim 26 wherein the solvent is ethanol or
isopropanol.
28. A process according to claim 18 wherein the solvent comprises ethanol
and from 1% to 5% water.
29. A process according to claim 18 wherein the amount of solvent used in
the extraction is from 1 to 9 times the volume of the oil to be treated.
30. An oil treated or prepared by a process according to claim 1.
31. An oil, comprising at least one polyunsaturated fatty acid (PUFA) that
has been produced by a microorganism, having a sterol content of no more
than 1.5%.
32. An oil according to claim 31 having a sterol content of no more than
1%.
33. An oil treated or prepared by a process according to claim 18.
34. An oil comprising at least one polyunsaturated fatty acid (PUFA)
produced by a microorganism, having a sterol content of at least 10%.
35. In a pharmaceutical, cosmetic, feed or foodstuff composition for
consumption by humans or animals having active ingredients wherein the
improvement comprises as an active ingredient the oil according to claim
30.
36. A composition comprising, or to which has been added, an oil according
to claim 30.
37. A composition according to claim 36 which is a foodstuff, feed,
cosmetic or pharmaceutical composition or a nutritional supplement for
consumption by humans or animals.
38. A composition according to claim 37 which is an infant formula.
39. In a pharmaceutical, cosmetic, feed or foodstuff composition for
consumption by humans or animals having active ingredients wherein the
improvement comprises as an active ingredient the oil according to claim
33.
40. A composition comprising, or to which has been added, an oil according
to claim 33.
41. A composition according to claim 40 which is a foodstuff, feed,
cosmetic or pharmaceutical composition or a nutritional supplement for
consumption by humans or animals.
42. A composition according to claim 41 which is an infant formula.
Description
FIELD OF THE INVENTION
The present invention relates to purified (such as by extraction)
polyunsaturated fatty acid (PUFA)-containing (microbial) oils, especially
oils with a triglyceride content of at least 97% and/or a sterol content
of either less than 1.5% or greater than 10%.
BACKGROUND OF THE INVENTION
There is a growing tendency to include lipid products containing
polyunsaturated fatty acids derived from various fermentation processes in
foodstuffs. This is of importance in the recently established desirability
to incorporate certain polyunsaturated fatty acids in an infant formula.
Various processes have been described for the fermentative production of
lipids or oils containing polyunsaturated fatty acids. Examples are
EP-A-155,420 for the production of .gamma.-linolenic acid (GLA)-containing
lipid from Mortierella, EP-A-223,960, EP-A-276,541 and WO-A-92/13086 for
the production of arachidonic acid (ARA)-containing oil from Mortierella
and/or Pythium, WO-A-91/07498 and WO-A-91/11918 for the production of
docosahexaenoic acid (DHA)-containing oil from Crypthecodinium cohnii or
Thraustochytrium, and WO-A-91/14427 for the production of eicosapentaenoic
acid (EPA)-containing oil from Nitzschia. Typically, the microbial species
producing the lipid containing the desired polyunsaturated fatty acid(s)
is cultured in a suitable medium and the biomass is harvested before the
desired lipid obtained.
To obtain a lipid concentrate which has a relatively high triglyceride
content typically a nonpolar solvent for the lipid (e.g. hexane) or
supercritical CO.sub.2 is used in the extraction process. For example,
EP-A-246,324 describes a fractional extraction process for the isolation
of lipids from Mortierella, to obtain different extracts which are
enriched in either polar or nonpolar (neutral) lipids. The neutral lipid
extract still has, however, a relatively low triglyceride content (89.3%)
and a high sterol content (9.4%). U.S. Pat. No. 4,857,329 describes an
extraction process comprising the use of supercritical CO.sub.2 to
selectively elute neutral lipids from Mortierella biomass. However, the
triglyceride content of the lipid extract does not exceed 86%.
Yamada et al, Industrial applications of single cell oils, Eds. Kyle and
Ratledge, 118-138 (1992) describe an arachidonic acid-containing oil
extracted from Mortierella alpina biomass using hexane. The purified oil
has a triglyceride content of 90%.
Thus, until now it has not been possible to obtain a microbial triglyceride
oil with a high triglyceride content, i.e. 95% or higher, using previous
fermentation and extraction technology. It has also not been possible to
prepare oils having a particularly low (e.g. less than 1.5%) or high (e.g.
at least 10%) sterol content.
DESCRIPTION OF THE INVENTION
The present invention generally relates to a process for preparing a
(microbial) oil with a high triglyceride content and a low content of
"unsaponifiables", where an oil extracted, obtained or derived from a
microbial biomass is treated with a polar solvent.
The present invention can thus provide a microbial (or microbially derived)
oil having a high triglyceride content, such as .gtoreq.95%. However the
oil may have a triglyceride content of at least 97%, preferably
.gtoreq.98%, and optimally .gtoreq.99%. The (microbial) oil may
alternatively or in addition have a low (e.g. .ltoreq.1.5%) or high (e.g.
.gtoreq.10%) sterol content. Preferably the sterol content is .ltoreq.1%,
such as .ltoreq.0.6%, optimally .ltoreq.0.3%.
The oil of the invention can be used in various compositions such as
pharmaceutical (or therapeutic), cosmetic, feedstuff or food compositions
(for human or animal consumption), especially in an infant formula or
nutritional supplement.
A first aspect of the present invention therefore relates to a process of
treating a microbially derived oil (an oil derived from a microorganism),
the process comprising contacting the oil with a polar solvent to extract
at least one compound that is soluble in the solvent, and separating at
least some of the solvent containing the compound from the (so treated)
oil.
The microbially derived oil can be extracted, obtained, or produced by one
or more microorganism(s). Often this will be the same species of
microorganism, but a mixture of two or more different microorganisms are
envisaged by the invention. The process of the invention may therefore be
subsequent to the production of the oil itself. The oil may be one that is
produced by, or exists inside (e.g. intracellularly) the microorganism(s).
Alternatively, it may be obtained from a (usually aqueous) composition
obtained or resulting from fermentation (of the microorganisms). This
(aqueous) composition may contain the microorganisms themselves: in that
case, it is usually a fermentation broth. The microorganisms (or biomass
as referred to in the art) can be removed (after fermentation) by a number
of methods, for example filtration, centrifugation or decantation. The oil
can be extracted or obtained from this biomass.
It is usual that the microbial oil will have been obtained by extraction.
This preferably will have involved extraction using a non-polar, or
preferably a water-immiscible, solvent, or at least a solvent that is
capable of extracting oily components. Such a solvent may be a C.sub.6-10
alkane, for example hexane, or (supercritical) carbon dioxide.
Different microorganisms will produce different oils. These can differ in
the amount of polyunsaturated fatty acids (PUFAs) as well as in other
components, and indeed the PUFAs may be in different forms, for example
diglycerides, triglycerides and/or phosolipids. As such, even microbially
derived oils can differ significantly from oils containing one or more of
these PUFAs that have been obtained from other (e.g. animal or fish or
vegetable) sources.
The microorganisms contemplated can vary widely, although preferably they
will be able to produce one or more PUFAs, for example on fermentation.
Microorganisms can be bacteria, algae, fungi or yeasts. Suitable
fermentation processes, microorganisms and PUFA-containing oils are
described in co-pending International application no. PCT/EP97/01448
(filed on Mar. 21, 1997 in the name of Gist-brocades B.V.), the content of
which is incorporated herein by reference.
Preferred algae are of the genus Crypthecodinium, Porphyridium or
Nitzschia. Preferred fungi are of the genus Thraustochytrium, Mortierella,
Pythium, Mucorales or Entomophthora, in particular of the species
Mortierella alpina.
The compound to be extracted can either be a desired compound, where the
compound is to be purified or even isolated, or it may be an impurity that
one wishes to remove from the oil. Generally speaking, the compound will
fall into the latter category. Thus, the compound may be an
"unsaponifiable", in other words one that is not solubilized (in water)
after treatment with an alkali (e.g. NaOH) and so does not form a salt
(thus it may not be capable of saponification). other compounds include
sterols, which can be alicyclic alcohols having a four conjugated ring
backbone, three aromatic C.sub.6 rings and one cyclopentane ring (e.g.
desmosterol, cholesterol) aliphatic and terpenic alcohols, tocopherol),
waxes and antifoaming agents, such as polypropylene glycol, which may be
present in the fermentation medium.
A second aspect of the present invention relates to a process of treating
an oil comprising at least one sterol, the process comprising contacting
the oil with a polar solvent to extract as least one sterol that is
soluble in the solvent, and separating at least some of the solvent
containing the sterol from the oil.
Preferred sterols include desmosterol, such as 5-desmosterol. If more than
one sterol is present, then suitably 70 to 90%, e.g. 80 to 85%, of the
sterols is desmosterol (e.g. for oil produced by Mortierella.
The oil will preferably contain at least one PUFA. This PUFA will usually
have been produced by the microbe or microorganism.
A third aspect of the invention relates to a process for preparing an oil
comprising at least one polyunsaturated fatty acid (PUFA), the process
comprising treating an oil comprising at least one PUFA and at least one
sterol with a polarsolvent to extract at least some of the PUFA and at
least some of the sterol (into the solvent), both the PUFA and the sterol
being at least partially soluble in the solvent, separating the solvent
(phase) from the oil (phase), and evaporating or otherwise removing some
of the solvent to give a (residual) oil having a sterol content of at
least 10%.
This sterol content may be even higher, such as at least 11%, for example
at least 14%.
PUFAs contemplated by the invention are C20 and C22 .omega.-3 and C18, C20
and C22 .omega.-6 polyunsaturated fatty acids. In particular they can
include .gamma.-linolenic acid (GLA), dihomo-.gamma.-linolenic acid (DLA),
arachidonic acid (ARA), eicosapentaenoic acid (EPA) and docosahexaenoic
acid (DHA). DHA is produced by algae or fungi, such as a dinoflagellate
algae, for example of the genus Crypthecodinium, or a fungus, for example
of the genus Thraustochytrium. GLA, DLA or ARA can be produced by fungi,
such as of the genus Mortierella, Pythium or Entomophthora. EPA can be
produced by an algae, such as of the genus Porphyridium or Nitzschia.
Typically the oil will dominantly or only contain one PUFA, although oils
can contain one or more PUFAs, for example in a lesser amount.
In the processes of the invention after the solvent has been added to the
oil, the two phases (oil and solvent) will usually separate. This can
easily then allow removal of one phase from the other.
It will be realised that in the second aspect of the invention one is
extracting a sterol from the oil. That can then give an oil with a low
sterol content, for example no more than 1.5%. The third aspect relates to
the processing of that solvent, in which some of the oil and sterol has
dissolved. That solvent will thus be relatively sterol rich: after some of
the solvent has been removed, one is left with a "residual" oil which can
have a sterol content of at least 10%.
A fourth aspect of the invention therefore relates to an oil treated or
prepared by a process according to any of the first to third aspects.
A fifth aspect relates to an oil, comprising at least one polyunsaturated
fatty acid that has been produced by a microorganism, having a sterol
content of no more than 1.5%. The (total) sterol content may in fact be no
more than 1%, for example less than 0.6%. By using the processes of the
invention, a sterol content of no more than 0.3% can be achieved.
A sixth aspect relates to an oil, comprising at least one polyunsaturated
fatty acid produced by a microorganism, having a sterol content of at
least 10%.
It will be realised that the oil of the fifth aspect can be prepared by
using the process of this second aspect, while the oil of the sixth aspect
can be prepared using the process of the third aspect.
The different oils of the invention can be prepared, for example, by using
different solvents, at different temperatures, as will be described later.
The present invention therefore provides a process for preparing an (e.g.
microbial) oil, where the oil is treated with one or more polar solvents.
These solvent(s) can therefore remove one or more compounds that are
soluble in the solvent. This may result in concentrating or enriching of
the oil. Therefore, if the oil contains triglycerides, one can concentrate
or increase the triglyceride content of the oil. This may be to at least
97%, for example at least 98%, and ultimately at least 99%.
Simultaneously with increasing the triglyceride content, the solvent
treatment can advantageously result in the removal of one or more
impurities from the oil. In particular, this treatment can result in the
lowering of the amount of "unsaponifiables". These unsaponifiables that
can be removed by the solvent treatment can include the sterols, aliphatic
and terpenic alcohols, waxes and antifoaming agents described earlier.
Usually, the treatment of the solvent will not alter the PUFA profile or
the oil so treated.
The polar solvent preferably comprises a C.sub.1-6 alkanol, for example
ethanol. The solvent, however, may be an aqueous one. Preferred solvents
therefore comprise an alcohol (e.g. ethanol) and water. However, the
solvent may comprise other liquids, and these can be acetone and/or
isopropanol.
If the solvent comprises ethanol, this may have a water content of from 0
to 20%, such as from 1 to 7%, and optionally from 2 to 4%. If the solvent
comprises methanol, acetone and/or isopropanol (IPA), then the water
content is preferably 0 to 2%, 5 to 50% and 5 to 15%, respectively. The
solvent may therefore comprise a mixture of two or more liquids. It has
been found that ethanol containing a small amount of water (e.g. 97%
ethanol, 3% water) can significantly improve the yield of triglyceride
after solvent treatment. This is because triglycerides are relatively
insoluble in this particular solvent. Having the solvent at a temperature
of from 15 to 30.degree. C., e.g. 20 to 25.degree. C., also reduces the
amount of triglycerides that dissolved in the solvent.
By using different solvents one can vary the amount of sterol (or indeed
PUFA) that is extracted. As has been discussed above, a mix of ethanol and
water can provide a high yield of triglycerides since although this
solvent will dissolve sterols, triglycerides are nevertheless relatively
insoluble in it.
The PUFA will generally exist in several forms, such as triglycerides and
diglycerides. These compounds are effectively a glycerol molecule with one
or more (although usually only one) of the PUFAs attached to this
backbone. Preferably the triglyceride form will be dominant. In the oil of
the fifth aspect (e.g. from the process of the second aspect), the amount
of diglycerides present is preferably no more than 2.2%, and preferably
less than 1%. The solvent used here is preferably at a temperature of from
10 to 40.degree. C., e.g. 20 to 30.degree. C.
In the oil of the sixth aspect, the relative ratios of triglycerides and
diglycerides can change. In the preparation of this oil, a solvent is
chosen to extract not only the sterol, but also some of the triglycerides
and diglycerides present in the original oil. The triglyceride content may
therefore vary from 60 to 90%. The diglyceride content may vary from 5 to
25, such as from 12 to 22%. It was found that ethanol with 3% water could
be a solvent for the diglycerides (and triglycerides) and so this solvent
is suitable for use in the process of the third aspect, for example to
produce an oil according to the sixth aspect. Here the solvent is
preferably employed at a temperature of from 50 to 70.degree. C., e.g. 55
to 65.degree. C.
The amount of compound to be extracted, or the triglyceride content, can be
adjusted by varying several process parameters. For example, one can
adjust the ratio of solvent to oil, the temperature during extraction
and/or by repeating the extraction process. If more than one extraction is
to be performed, a counter-current extraction process is preferred, which
can minimise triglyceride losses.
Usually the oil will be a crude oil obtained after extraction from a (e.g.
dried) microbial biomass with a suitable solvent, followed by evaporation
of that (water immiscible) solvent. The oil may be subjected to one or
more refining steps prior to the process of the invention.
The oil of the invention, or one which results from a process of the first,
second or third aspect, can be used for various purposes without further
processing, or can be additionally subjected to one or more refining
steps. The oil can be used as an additive or a supplement, for example in
food compositions, such as an infant formula. It may however also be used
in cosmetic or pharmaceutical compositions. The invention in a further
aspect therefore relates to a composition, such as a food stuff, feed or
pharmaceutical composition or a cosmetic composition, which comprises, or
to which has been added, an oil of the invention. Preferred compositions
are foods, such as infant formula or a nutritional supplement.
The oil of the invention can therefore have a low sterol and/or low
diglyceride content. It may also have a high triglyceride content. This
makes the oil particularly suitable for nutritional purposes, and can be
used as a nutritional supplement. The oil may be supplied as an oil, or it
may be encapsulated, for example, in a gelatin capsule. The oil can thus
be incorporated in foods, feeds or foodstuffs, suitable for human or
animal consumption. Suitable examples are health drinks and bread.
Particularly contemplated is the use in infant formula, or in cosmetics.
Preferred features and characteristics of one aspect of the invention are
equally applicable to another aspect mutatis mutandis.
The invention will now be described, by way of example, with reference to
the following Examples which are provided merely for means of
illustration, and are not to be construed as being limiting on the
invention.
COMPARATIVE EXAMPLE 1
Recovery of crude ARA oil from M. alpina biomass
500 l of broth obtained after Mortierella alpina fermentation was filtered
in a membrane filter press (cloth type: propex 46K2). The broth was
filtered with a pressure difference of 0.2 bar. Within 21 minutes 500 l
broth was filtered over a total filter area of 6.3 m.sup.2 which resulted
in an average flow of about 230 l/m.sup.2 h. The filter cake was washed in
30 minutes with 10 cake volumes of tap water at an average flow rate of
320 l/m.sup.2 h.
The cake was squeezed at 5.5 bar for 30 minutes which resulted in a dry
matter content of the recovered biomass of about 45%.
Extrusion was performed on the resulting biomass cake using a single screw
extruder with a profiled barrel and a universal screw. The dieplate used
for extrusion had holes of diameter 2 mm.
Drying of the extrudate was performed in a fluidized bed dryer with air
(8000 Nm.sup.3 /m.sup.2 h). The setpoint of the bed temperature was
80.degree. C. The diameter of the dried extruded biomass was 2 mm and its
dry matter content after drying was about 96%.
A crude arachidonic acid-containing oil (ARA oil) was then extracted from
the extrudate using hexane as a solvent.
EXAMPLES 2 AND 3
Treatment of microbial ARA oil with 100% ethanol
5 ml of crude ARA oil was extracted from the extrudate of Example 1 with a
volume of 100w ethanol for 1 minute by hand-shaking. Subsequently, the
bottom and toplayers were separated by centrifugation for 5 minutes at
5000 rpm. The samples were analyzed by means of (600 Mhz) NMR (for tri-
and di-glycerides, sterols (only desmosterol content measured) and
antifoaming agent).
Extraction of crude ARA oil with 9 volumes of 100t ethanol at two different
temperatures resulted in an oil with a decreased level of sterol and
diglyceride (DG) and in an increased level of triglyceride (TG, see Table
1). The yield of TG is the percentage of triglyceride remaining in the oil
after solvent extraction. Also antifoaming agent was removed and found in
the ethanol after extraction. However, the yield of triglycerides was low
due to the fact that some of the TG dissolved (and was thus removed in)
the ethanol.
TABLE 1
______________________________________
Extraction of crude ARA oil with 100% ethanol
(data for treated oil)
Ex solvent temp. % TG % DG % sterol
yield TG
______________________________________
-- Control -- 96.2 2.2 1.6 100
2 EtOH 100% ambient 98.2 0.7 1.1 73.8
3 EtOH 100% 60.degree. C. 98.5 0.7 0.8 43.2
______________________________________
Key:
TG: triglycerides
DG: diglycerides
Sterol: as desmosterol
EXAMPLES 4 TO 9
Treatment of microbial ARA oil with 97% ethanol
Examples 2 and 3 were repeated except using 97% ethanol at varying volumes
relative to the oil.
Extraction of crude ARA oil with 1, 3 and 9 volumes of 97% ethanol resulted
in an oil with a decreased level of sterol and diglyceride and in an
increased level of triglyceride (see Table 2).
The yield of triglycerides was above 92% due to the fact that not much oil
dissolves in 97% ethanol. At ambient temperature (about 20.degree. C.), a
higher yield of triglycerides and a better removal of diglycerides and
sterols was observed. Remarkably no ethanol was found in the treated oil.
TABLE 2
______________________________________
Extraction of crude ARA oil with 97% ethanol
(data for treated oil)
vol %
Ex solvent temp. EtOH % TG % DG sterol yield TG
______________________________________
-- Control -- 0 96.2 2.2 1.6 100
4 EtOH 97% ambient 1 96.7 1.8 1.4 92.9
5 EtOH 97% ambient 3 97.8 1.1 1.1 95.0
6 EtOH 97% ambient 9 98.9 0.4 0.7 96.2
7 EtOH 97% 60.degree. C. 1 96.4 2.0 1.6 99.7*
8 EtOH 97% 60.degree. C. 3 97.7 1.1 1.2 92.4
9 EtOH 97% 60.degree. C. 9 98.3 0.6 1.1 93.7
______________________________________
Key:
TG: triglycerides
DG: diglycerides
Sterol: as desmosterol
*Due to the increase of the lower (oil) phase because the ethanol partly
dissolved into the oil and so phase separation was more difficult.
The ethanol phase was also analyzed after extraction and a significant
increase in sterols was observed. Also the antifoam agent (polypropylene
glycol) was extracted and found in the ethanol phase (see Table 3).
TABLE 3
______________________________________
Extraction of crude ARA oil with 97% ethanol
(data for ethanol phase)
vol % %
Ex solvent temp. EtOH % TG % DG antifoam sterol
______________________________________
4 EtOH 97% ambient 1 60.9 20.8 4.1 14.2
5 EtOH 97% ambient 3 73.1 15.3 1.3 10.2
6 EtOH 97% ambient 9 83.0 10.0 0.7 6.3
7 EtOH 97% 60.degree. C. 1 66.1 18.3 3.7 11.9
8 EtOH 97% 60.degree. C. 3 78.6 12.5 1.1 7.8
9 EtOH 97% 60.degree. C. 9 87.9 7.1 0.4 4.5
______________________________________
Key:
TG: triglycerides
DG: diglycerides
Sterol: as desmosterol
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